Sera from halothane hepatitis patients have been shown to contain antibodies that react with several trifluoroacetylated proteins (100 kDa, 76 kDa, 59 kDa, 57 kDa, 54 kDa) purified from the livers of halothane treated rats. These findings suggest that similar trifluoroacetylated proteins are the immunogens responsible for the formation of the patient's antibodies and raise the possibility that their formation may result in hepatitis. Recently, the 59 kDa protein has been identified as a carboxylesterase. In order to further characterize this protein, we have begun to clone it. Polyclonal anti-59 kDa antibodies were raised in rabbits and were used for screening cDNA libraries constructed in the expression vector lambda gt11. Several positive clones were isolated. A cDNA from a rat liver library containing a complete protein reading frame and putative active site regions for a carboxylesterase was completely sequenced. Clones obtained from a human liver library are currently being analyzed and should yield complete genetic and protein sequence data for a human carboxylesterase isoenzyme. These clones will be used in the future to determine the distribution, regulation, and physiological function of the liver microsomal carboxylesterases and to elucidate the immunodominant domains and role in halothane hepatitis of the trifluoroacetylated 59 kDa carboxylesterase.